3-1 Cyclin-Dependent Kinases

The cyclin-dependent kinases are a small family of enzymes that

require cyclin subunits for activity

The cyclin-dependent kinases (Cdks) are a family of serine/threonine protein kinases whose
members are small proteins (~3440 kDa) composed of little more than the catalytic core
shared by all protein kinases. By definition, all Cdks share the feature that their enzymatic
activation requires the binding of a regulatory cyclin subunit. In most cases, full activation also
requires phosphorylation of a threonine residue near the kinase active site.
Although originally identified as enzymes that control cell-cycle events, members of the Cdk
family are involved in other cellular processes as well. Animal cells, for example, contain at least
nine Cdks, only four of which (Cdk1, 2, 4 and 6) are involved directly in cell-cycle control
(Figure 3-2). Another family member (Cdk7) contributes indirectly by acting as a Cdk-activating
kinase (CAK) that phosphorylates other Cdks, as we discuss in section 3-3. Cdks are also
components of the machinery that controls basal gene transcription by RNA polymerase II
(Cdk7, 8 and 9) and are involved in controlling the differentiation of nerve cells (Cdk5).
We will focus on the small number of Cdks for which there is clear evidence of a direct role
in cell-cycle control (see Figure 3-2). In the fission yeast Schizosaccharomyces pombe and the
budding yeast Saccharomyces cerevisiae (see section 2-1), all cell-cycle events are controlled by a
single essential Cdk called Cdk1. Cell-cycle events in multicellular eukaryotes are controlled
by two Cdks, known as Cdk1 and Cdk2, which operate primarily in M phase and S phase,
respectively. Animal cells also contain two Cdks (Cdk4 and Cdk6) that are important in
regulating entry into the cell cycle in response to extracellular factors.
Cdk function has been remarkably well conserved during evolution. It is possible, for example,
for yeast cells to proliferate normally when their gene for Cdk1 is replaced with the human
one. This and other evidence clearly illustrates that Cdk function, and thus the function of the
cell-cycle control system, has remained fundamentally unchanged over hundreds of millions of
years of eukaryotic evolution.
Cyclin-Dependent Kinases
Species

Name

Size
Original (amino
name acids)

Function

S. cerevisiae

Cdk1

Cdc28

298

all cell-cycle
stages

S. pombe

Cdk1

Cdc2

297

all cell-cycle
stages

D. melanogaster

Cdk1

Cdc2

297

Cdk2

Cdc2c

314

G1/S, S,
possibly M

Cdk4

Cdk4/6

317

G1
promotes growth

Cdk1

Cdc2

301

297

S,
possibly M

X. laevis

Cdk2

H. sapiens

Cdk1

297

Cdk2

Cdc2

298

G1/S, S,
possibly M

Cdk4

303

G1

Cdk6

326

G1

Definitions
L12 helix: small alpha helix adjacent to the T-loop in
the active site of Cdk2 (residues 147151), which
changes structure to a beta strand upon cyclin binding.

Cdks exert their effects on cell-cycle events by phosphorylating a large number of proteins in
the cell. During mitosis in particular, when many aspects of cellular architecture and metabolism
are altered, Cdks phosphorylate hundreds of distinct proteins. These Cdk substrates are
phosphorylated at serine or threonine residues in a specific sequence context that is recognized
by the active site of the Cdk protein. In most cases, the target serine (S) or threonine (T)
residue is followed by a proline (P); it is also highly favorable for the target residue to have a
basic amino acid two positions after the target residue. The typical phosphorylation sequence
for Cdks is [S/T*]PX[K/R], where S/T* indicates the phosphorylated serine or threonine, X
represents any amino acid and K/R represents the basic amino acid lysine (K) or arginine (R).
The active site of cyclin-dependent kinases is blocked in the absence of
cyclin

All protein kinases have a tertiary structure comprising a small amino-terminal lobe and a
larger carboxy-terminal lobe. ATP fits snugly in the cleft between the lobes, in such a way that
the phosphates are oriented outwards, toward the mouth of the cleft. The protein substrate
Figure 3-2 Table of cyclin-dependent kinases that control the cell cycle

T-loop: flexible loop adjacent to the active site of Cdks,

named for the threonine whose phosphorylation is
required for maximal activity. Sometimes called the
activation loop.

PSTAIRE helix: alpha helix in the amino-terminal lobe

of Cdks (also known as the a1 helix), which interacts
with cyclin and is moved inward upon cyclin binding,
resulting in reorientation of key active-site residues.The
name of this helix comes from its amino-acid sequence,
which is conserved among all major Cdks.

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2007 New Science Press Ltd

Cyclin-Dependent Kinases 3-1

binds at the entrance of the cleft, interacting mainly with the surface of the carboxy-terminal
lobe. Nearby residues catalyze the transfer of the terminal g-phosphate of ATP to a hydroxyl
oxygen in the protein substrate.
Cdks have the same two-lobed structure as other protein kinases (Figure 3-3), but with two
modifications that make them inactive in the absence of cyclin. These modifications have been
revealed by detailed crystallographic studies of the structure of human Cdk2. First, a large,
flexible loopthe T-loop or activation looprises from the carboxy-terminal lobe to block
the binding of protein substrate at the entrance of the active-site cleft. Second, in the inactive
Cdk several important amino-acid side chains in the active site are incorrectly positioned, so
that the phosphates of ATP are not ideally oriented for the kinase reaction. Cdk activation
therefore requires extensive structural changes in the Cdk active site.
Two alpha helices make a particularly important contribution to the control of Cdk activity.
The highly conserved PSTAIRE helix of the upper kinase lobe (also known as the a1 helix)
interacts directly with cyclin and moves inward upon cyclin binding, causing the reorientation
of residues that interact with the phosphates of ATP. The small L12 helix, just before the T-loop
in the primary sequence, changes structure to become a beta strand upon cyclin binding, also
contributing to reconfiguration of the active site and T-loop. We discuss the structural basis
of Cdk activation in more detail in section 3-4. First, we will describe the cyclins and other
regulators that influence activation.

(b)

(a)

N
1

HsCdk2
HsCdk1
SpCdk1
ScCdk1

MENFQKVEKIGEGTYGVVYKARNK LTGE VVALKKIRLDTETEGVPSTAIREISLLKELN

HPNIVKLLD
MEDYTKIEKIGEGTYGVVYKGRHK TTGQ VVAMKKIRLESEEEGVPSTAIREISLLKELR
HPNIVSLQD
MENYQKVEKIGEGTYGVVYKARHK LSGR IVAMKKIRLEDESEGVPSTAIREISLLKEVNDENNRSNCVRLLD
MSGELANYKRLEKVGEGTYGVVYKALDLRPGQGQRVVALKKIRLESEDEGVPSTAIREISLLKELK
DDNIVRLYD

HsCdk2
HsCdk1
SpCdk1
ScCdk1

VIHTE NKLYLVFEFLHQDLKKFMDASAL
TGIPLPLIKSYLFQLLQGLAFCHSHRVLHRDLKPQNLLINTEGAIK
VLMQD SRLYLIFEFLSMDLKKYLDSIPP GQYMDSSLVKSYLYQILQGIVFCHSRRVLHRDLKPQNLLIDDKGTIK
ILHAE SKLYLVFEFLDMDLKKYMDRISETGATSLDPRLVQKFTYQLVNGVNFCHSRRIIHRDLKPQNLLIDKEGNLK
IVHSDAHKLYLVFEFLDLDLKRYMEGIPK DQPLGADIVKKFMMQLCKGIAYCHSHRILHRDLKPQNLLINKDGNLK

L12

T-loop

PSTAIRE
helix

142
143
149
151

helix L12

LADFGLARAFGVPVRTYTHEVVTLWYRAPEILLGCKYYSTAVDIWSLGCIFAEMVTRRALFPGDSEIDQLFRIFRTLG
LADFGLARAFGIPIRVYTHEVVTLWYRSPEVLLGSARYSTPVDIWSIGTIFAELATKKPLFHGDSEIDQLFRIFRALG
LADFGLARSFGVPLRNYTHEIVTLWYRAPEVLLGSRHYSTGVDIWSVGCIFAEMIRRSPLFPGDSEIDEIFKIFQVLG
LGDFGLARAFGVPLRAYTHEIVTLWYRAPEVLLGGKQYSTGVDTWSIGCIFAEMCNRKPIFSGDSEIDQIFKIFRVLG

HsCdk2
HsCdk1
SpCdk1
ScCdk1

TPDEVVWPGVTSMPDYKPSFPKWARQDFSKVVPPLDEDGRSLLSQMLHYDPNKRISAKAALAHPFFQDVTKPVPHLRL
TPNNEVWPEVESLQDYKNTFPKWKPGSLASHVKNLDENGLDLLSKMLIYDPAKRISGKMALNHPYFNDLDNQIKKM
TPNEEVWPGVTLLQDYKSTFPRWKRMDLHKVVPNGEEDAIELLSAMLVYDPAHRISAKRALQQNYLRDFH
TPNEAIWPDIVYLPDFKPSFPQWRRKDLSQVVPSLDPRGIDLLDKLLAYDPINRISARRAAIHPYFQES

HsCdk2
HsCdk1
SpCdk1
ScCdk1

L14

68
68
72
75

220
221
227
229

7
298
297
297
298

Figure 3-3 Cyclin-dependent kinase structure (a) Amino-acid sequences of major Cdks controlling the cell cycle in humans (H. sapiens (Hs) Cdk1 and Cdk2)
and yeast (S. pombe (Sp) Cdk1 and S. cerevisiae (Sc) Cdk1). Yellow residues are identical in all four kinases. Above the alignment, secondary structure
elements in human Cdk2 are shown for comparison with the tertiary structure in panel (b). Key landmarks are highlighted, including the PSTAIRE or a1
helix, the inhibitory phosphorylation sites Thr 14 and Tyr 15, the activating phosphorylation site (Thr 160 in human Cdk2) and the T-loop or activation loop
where Thr 160 is found. (b) Tertiary structure of human Cdk2, determined by X-ray crystallography. Like other protein kinases, Cdk2 is composed of two
lobes: a smaller amino-terminal lobe (top) that is composed primarily of beta sheet and the PSTAIRE helix, and a large carboxy-terminal lobe (bottom) that is
primarily made up of alpha helices. The ATP substrate is shown as a ball-and-stick model, located deep within the active-site cleft between the two lobes.
The phosphates are oriented outward, toward the mouth of the cleft, which is blocked in this structure by the T-loop (highlighted in green). (PDB 1hck)

References
De Bondt, H.L. et al.: Crystal structure of cyclindependent kinase 2. Nature 1993, 363:595602.
Morgan, D.O.: Cyclin-dependent kinases: engines,
clocks, and microprocessors. Annu. Rev. Cell Dev. Biol.
1997, 13:261291.
Ubersax, J.A. et al.: Targets of the cyclin-dependent
kinase Cdk1. Nature 2003, 425:859864.

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